Tumor markers for human tumor cells have been largely limited to activated oncogenes and their products, for example, the myc, ras, fos, and erbB2 genes and their encoded oncoproteins. In addition, activated anti-oncogenes, such as RB, p53, and DCC, have been identified in normal cells but do not appear to be present in tumor cells. Oncogene and anti-oncogene products have proven difficult to use as consistent predictors of tumor and normal tissue, respectively, due to the relatively low level of expression of the genes encoding these proteins. Thus, there is a need in the art for a tumor marker which is not only differentially expressed in tumor and normal tissue, but also consistently detectable in human tumor tissue and consistently absent in the corresponding normal tissue.
A common method used to identify genes differentially or uniquely expressed in tumors, in cells responding to growth factors, and in differentiated cell types such as, among others, T cells, adipocytes, neurons, and hepatocytes is the subtractive hybridization technique (S. W. Lee et al., Proc. Natl. Acad. Sci. USA 80:4699, 1983). A method of differential display of eukaryotic mRNA by means of the polymerase chain reaction (PCR) has recently been developed (P. Liang et al., Science 257:967, 1992). This method utilizes oligo dT linked to two additional bases as the primer for reverse transcription driven by reverse transcriptase. cDNA fragments are then amplified by Taq DNA polymerase-based PCR using an oligo dT primer along with one additional primer. The amplified cDNAs are then resolved by DNA sequencing gels. There is a need in the art for a streamlined and simplified process for isolating cDNAs corresponding to differentially expressed mRNAs.